More than 70 years after the first ex situ genebanks have been established, major efforts in this field are still concerned with issues related to further completion of individual collections and securing of their storage. Attempts regarding valorization of ex situ collections for plant breeders have been hampered by the limited availability of phenotypic and genotypic information. With the advent of molecular marker technologies first efforts were made to fingerprint genebank accessions, albeit on a very small scale and mostly based on inadequate DNA marker systems. Advances in DNA sequencing technology and the development of high-throughput systems for multiparallel interrogation of thousands of single nucleotide polymorphisms (SNPs) now provide a suite of technological platforms facilitating the analysis of several hundred of Gigabases per day using state-of-the-art sequencing technology or, at the same time, of thousands of SNPs. The present review summarizes recent developments regarding the deployment of these technologies for the analysis of plant genetic resources, in order to identify patterns of genetic diversity, map quantitative traits and mine novel alleles from the vast amount of genetic resources maintained in genebanks around the world. It also refers to the various shortcomings and bottlenecks that need to be overcome to leverage the full potential of high-throughput DNA analysis for the targeted utilization of plant genetic resources.